Abstract
Human small heat shock protein HspB6 (Hsp20) was modified by metabolic α-dicarbonyl compound methylglyoxal (MGO). At low MGO/HspB6 molar ratio, Arg13, Arg14, Arg27, and Arg102 were the primary sites of MGO modification. At high MGO/HspB6 ratio, practically, all Arg and Lys residues of HspB6 were modified. Both mild and extensive MGO modification decreased susceptibility of HspB6 to trypsinolysis and prevented its heat-induced aggregation. Modification by MGO was accompanied by formation of small quantities of chemically crosslinked dimers and did not dramatically affect quaternary structure of HspB6. Mild modification by MGO did not affect whereas extensive modification decreased interaction of HspB6 with HspB1. Phosphorylation of HspB6 by cyclic adenosine monophosphate (cAMP)-dependent protein kinase was inhibited after mild modification and completely prevented after extensive modification by MGO. Chaperone-like activity of HspB6 measured with subfragment 1 of skeletal myosin was enhanced after MGO modifications. It is concluded that Arg residues located in the N-terminal domain of HspB6 are easily accessible to MGO modification and that even mild modification by MGO affects susceptibility to trypsinolysis, phosphorylation by cAMP-dependent protein kinase, and chaperone-like activity of HspB6.
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Abbreviations
- DTT:
-
Dithiotreitol
- MDH:
-
Malate dehydrogenase
- ME:
-
β-Mercaptoethanol
- MGO:
-
Methylglyoxal
- PMSF:
-
Phenylmethanesulfonyl fluoride
- S1:
-
Subfragment 1 of rabbit skeletal muscle myosin
- SEC:
-
Size-exclusion chromatography
- TPCK:
-
Nα-tosyl-l-phenylalanine chloromethyl ketone
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Acknowledgments
Investigation of physico-chemical properties was supported by the Russian Science Foundation (RSF) (grant no. 14-35-00026). Investigation of protein-protein interaction was supported by the Russian Foundation for Basic Research (grant no. 16-04-00016). MALDI MS facility became available in the framework of the Moscow State University Development Program PNG 5.13. The authors are grateful to Dr. O.P. Nikolaeva and Prof. D.I. Levitsky (A.N. Bach Institute of Biochemistry, Russian Academy of Sciences) for providing subfragment-1 of rabbit skeletal myosin.
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Muranova, L.K., Perfilov, M.M., Serebryakova, M.V. et al. Effect of methylglyoxal modification on the structure and properties of human small heat shock protein HspB6 (Hsp20). Cell Stress and Chaperones 21, 617–629 (2016). https://doi.org/10.1007/s12192-016-0686-4
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DOI: https://doi.org/10.1007/s12192-016-0686-4